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Technical Abstract: Soybean is an important crop which requires additional tools for advanced genetic manipulation of valuable traits. Soybean oil typically consists of undesirably high amounts of the oxidatively- unstable linolenic acid component of the total fatty acids. Breeding for low linolenic acid soybeans is possible with mutagenized parent lines, but it is complicated by the complex nature of the trait. The objective of this research was to develop an understanding of the molecular genetics controlling soybean seed linolenic acid levels. Using a candidate-gene approach based on the Arabidopsis thaliana microsomal omega-3 fatty acid desaturase sequence (FAD3), we identified three soybean FAD3 homologues. In low linolenic mutant soybean lines, deleterious mutations were identified in the homologous genes. Molecular markers were devised to distinguish mutant alleles. In segregating F2 populations, replacement of wild-type alleles with the corresponding mutant alleles resulted in an unequal stepwise reduction in linolenic acid levels. The mutant genotype conferred the low linolenic acid phenotype in subsequent generations. We are building a model to describe the contribution of each of the three soybean FAD3 genes to the seed linolenic acid levels so soybean breeders can determine the best combination of mutant alleles to generate desired fatty acid profiles.